Cystic fibrosis (CF) is a recessive genetic disease caused by a single defective gene located on the long arm of chromosome 7. The carrier frequency in the U.S. Caucasian population is about 1 in 23. The clinical syndrome includes elevated sweat electrolytes, pancreatic insufficiency, thickened cervical mucus in females and blockage and eventual degeneration of the vas deferens in males, susceptibility to meconium ileus in newborns and an equivalent form of intestinal blockage later, and chronic lung infection, usually by gram negative bacteria. Eventually complications from the lung infections lead to death; the median survival age in the U.S. is now about 21 years. We propose to help understand CF by using electrophysiological techniques to localize the relevant gene product, clarify how it works, and determine where it is expressed. We take as our starting point that the gene product helps regulate a C1- ion channel complex that is a portion of cAMP-regulated ion transport. The defective gene product is expressed in a variety of exocrine cells, where it results in reduced fluid secretion and reduced salt absorption. The proposed experiments are designed to fill in some of the many gaps between the defective gene and the clinical syndrome. The experiments are designed to decide between two possibilities: Is the defective gene product inextricably linked to C1- ion conductance, or is C1- ion conductance simply one of several mechanisms affected by a defect in a more general regulatory product? Our specific aims are (1) to analyze apical chloride channel gating in cultured cells using patch-clamp techniques; and to determine: (2) if the CF gene is expressed by heterozygotes; (3) if the reabsorption of NaC1 by sweat ducts is regulated by a cAMP-dependent increase in chloride conductance; and (4) if beta-adrenergically stimulated sweating depends upon regulation of chloride channels by a cAMP-dependent pathway. We also propose to establish the CF phenotype in cultured cell types not presently studied. For each of the systems we study, we plan to begin to trace the consequences of decreased C1- conductance on cell and organ system function. In addition to on-cell, whole-cell and inside-out excised patch-clamp techniques, we will use intracellular microelectrode recording and microperfusion of sweat ducts.